My research project is focused mainly on the characterization of the active site structure of the mammalian hemoprotein peroxidase, myeloperoxidase, which catalyzes the one-electron oxidation of micro- and macromolecular substrates. The three-dimensional structure of myeloperoxidase has been solved to 2.28 A. This enzyme is a tetramer composed of two heavy subunits of Mr 57000-60000 and two light subunits of Mr 10000-15000. A single disulfide bridge links the two heavy subunits, which each binds a heme group. Myeloperoxidase is found primarily in azurophil granules of mammalian neutrophils where it catalyzes the hydrogen peroxide-mediated peroxidation of chloride ion to hypochlorite, which with its secondary metabolites is effective in killing phagocytized bacteria and viruses. The enzyme might also have a role in aggravating tissue degradation in inflammatory diseases. The heme groups in myeloperoxidase appear to be covalently bound to the enzyme, although there is conflicting evidence as to the exact nature and location of these attachments, despite the solution of the X-ray crystal structure. I will attempt to identify the amino acid residues involved in the catalytic mechanism and heme binding of myeloperoxidase through site-directed mutagenesis and kinetic and spectroscopic characterizations of expressed mutant proteins. I will also explore in detail the exact nature of the putative covalent linkages between the enzyme and its heme groups. In order to accomplish these goals, I need access to modelling programs in the Computer Graphics Laboratory that will allow me to examine the active site structures of peroxidases whose crystal structures are known. I also need to perform DNA and protein sequence comparisons with a sequence comparison program and have access to sequence data banks.